Project Abstract Fragile X Syndrome (FXS) is a leading inheritable cause of mental impairment. There is no known cure for FXS or treatment that reverses the collective pathology. There is a fundamental gap in our knowledge of how FXS causes mental impairments through alteration of neural circuitry. The long-term goal of this research is to develop an understanding of FXS that links learning impairments to specific changes in neural circuits. Characteristic symptoms of FXS include reduced intellectual abilities, learning deficits, and hypersensitivity to sensory stimuli. FXS arises from a loss-of-function in the FMR1 gene; mice lacking a functional fmr1 gene exhibit several phenotypes similar to FXS. Fmr1 mutant mice display an intriguing deficit on both the gap cross task, a freely-behaving whisker-dependent tactile learning task, as well as a head-fixed whisker-dependent tactile learning task. The central hypothesis is that impairments in tactile learning are driven by reduced dendritic spine stability and hypersensitive touch responses in primary somatosensory cortex resulting from attenuated activity of somatostatin-expressing (SOM) interneurons. Experiments in this proposal will determine the extent to which loss of the fmr1 gene disrupts spine stability, tactile learning, and circuit dynamics during task performance. Guided by our strong preliminary data, we will pursue this hypothesis in two related specific aims. In Aim 1, longitudinal two-photon in vivo imaging is combined with an automated head-fixed whisker- dependent tactile learning task to evaluate if reduced activity of SOM interneurons in fmr1 mutant mice decreases dendritic spine stability and impairs learning. In Aim 2, sophisticated electrophysiology is combined with high-speed tracking of whisker position during this same head-fixed object localization to quantify the extent to which tactile discrimination and cortical representations of afferent sensory activity in somatosensory cortex are abnormal fmr1 mutant mice and if attenuated function of SOM interneurons contributes to this deficit. This approach is particularly innovative because the synaptic changes that underlie learning are measured longitudinally throughout task acquisition. Furthermore, breaking from the anesthetized status quo, the cortical circuit dynamics that represent touch are quantified during active perceptual behavior. The proposal is significant because it vertically advances our knowledge of FXS mechanisms across levels of analysis, from synapse to circuit to behavior. Additionally, it opens new horizons for these advanced techniques to be applied to other cortical layers and brain regions to build a comprehensive understanding of neural circuit defects in a premier FXS model system. This proposal squarely meets the key mission objectives of the NINDS and NIMH to provide detailed and integrated knowledge of how the function of synapses and circuits is disrupted in neurological disorders. Ultimately, the resulting improved understanding of circuit dysfunction has the potential to lead to therapies that improve the quality of life for the roughly 1 in 5,000 people born with Fragile X Syndrome.